Generation of formic acid and ethanolamine from serine in biosynthesis of linear gramicidin by a cell-free preparation of Bacillusbrevis (ATCC 8185)

A growing organism that produces antibiotic peptide was incubated with L-(U-¹⁴C)serine for labeling linear gramicidin. Linear gramicidin was isolated by a simple chromatographic method from tyrothricin (mixture of linear gramicidin and tyrocidine) applied to a column of basic aluminum oxide. The hydrolysate of labeled linear gramicidin on thin layer chromatography showed that L-(U-¹⁴C)serine was one of a precursor of ethanolamine moiety by autoradiography. L-(3-¹⁴C)serine generated formic acid in the presence of tetrahydrofolic acid by an enzyme fraction prepared with ammonium sulfate, and further formed ethanolamine binding to the protein. Formylvaline was biosynthesized by it with tetrahydrofolic acid and ATP, and subsequently released from the protein.     

Maturation and fertilization of porcine oocytes in vitro

Four experiments were conducted to study 1) factors affecting porcine oocyte maturation in culture medium and 2) a new method for oocyte maturation outside the porcine body. In Experiment 1, five groups of oocytes were cultured in m-TCM199 or m-KRB medium for 24 to 28, 32 to 36 or 40 to 42 hours and then were fertilized in vitro. The cleavage rate (two to four-cell stage) of oocytes cultured for 32 to 36 hours was significantly higher than those of the other oocytes. The results indicate that a suitable culture period for the in vitro maturation of porcine oocytes is 32 to 36 hours. In Experiment 2, four groups of oocytes were cultured in m-KRB or m-KRB supplemented with PFF, PMSG or FSH for in vitro maturation, and the cleavage rates of oocytes were 7.94, 22.56, 30.23 and 23.26%, respectively, after in vitro fertilization. The results show that porcine follicular fluid (PFF) and gonadotrophins added to the culture medium promote porcine oocyte maturation in vitro. In Experiment 3, oocytes were cultured in m-KRB or m-TCM199, supplemented with both gonadotrophin and pocine folliclar fluid for maturation in vitro. After fertilization in vitro, the cleavage rates of oocytes were 26.32 and 27.93% for the two media. The results indicate that the difference between m-KRB and m-TCM199 was insignificant when the media were used to culture porcine oocytes. But there was a significant difference when PFF and gonadotrophins were added to the basic media. In Experiment 4, porcine oocytes were transferred into the reproductive tracts of other animals for maturation. After 34 to 36 hours, the oocytes were collected and fertilized in vitro. The cleavage rates of oocytes were 10.42, 28.45, 3.33 and 36.36%, respectively, for the oocytes matured in mouse uterine horns, rat uterine horns, rat oviducts or rabbit oviducts. The results show that porcine oocytes can be matured in the reproductive tracts of other animals.     

基于科研素质培养的“分子生物学”课程教学改革

作为迅速发展的前沿学科及连续性与承接性很强的实验性学科,"分子生物学"课程对于构建研究生知识体系、培养研究生的科研素养、创新意识和工程实践能力具有重要作用。为满足"新工科"人才培养的需求,我们基于成果导向教育(Outcome-Based Education,OBE)理念,整合教学内容,改革教学方法,借助现代信息化教学平台,以学习小组为单位,开展课堂竞猜、翻转课堂、课堂讨论等教学活动;运用基于问题的学习(Problem-BasedLearning,PBL)法,建立"互动式"的教学模式;设计过程化考核与终结性考核相结合的多元化考核体系,培养研究生的创新能力和工程实践能力,建立一种适合于本校生物工程专业研究生培养的"分子生物学"教学体系。研究生参与教学效果评估及科研成果随访的结果表明,该探索取得了良好的教学效果,为"新工科"背景下生物工程领域的研究型综合创新人才的培养奠定了基础。     

Systematic Evaluation of Protein Sequence Filtering Algorithms for Proteoform Identification Using Top‐Down Mass Spectrometry

Complex proteoforms contain various primary structural alterations resulting from variations in genes, RNA, and proteins. Top‐down mass spectrometry is commonly used for analyzing complex proteoforms because it provides whole sequence information of the proteoforms. Proteoform identification by top‐down mass spectral database search is a challenging computational problem because the types and/or locations of some alterations in target proteoforms are in general unknown. Although spectral alignment and mass graph alignment algorithms have been proposed for identifying proteoforms with unknown alterations, they are extremely slow to align millions of spectra against tens of thousands of protein sequences in high throughput proteome level analyses. Many software tools in this area combine efficient protein sequence filtering algorithms and spectral alignment algorithms to speed up database search. As a result, the performance of these tools heavily relies on the sensitivity and efficiency of their filtering algorithms. Here, we propose two efficient approximate spectrum‐based filtering algorithms for proteoform identification. We evaluated the performances of the proposed algorithms and four existing ones on simulated and real top‐down mass spectrometry data sets. Experiments showed that the proposed algorithms outperformed the existing ones for complex proteoform identification. In addition, combining the proposed filtering algorithms and mass graph alignment algorithms identified many proteoforms missed by ProSightPC in proteome‐level proteoform analyses.     

RETRACTED ARTICLE: Therapeutic DNA Vaccination as a Repair Strategy Following Spinal Cord Injury

Myelin-derived proteins, such as tenascin-R (TN-R), myelin associate glycoprotein (MAG), oligodendrocyte-myelin glycoprotein (OMgp), and Nogo-A, inhibit the central nervous system regeneration. In this study, the DNA vaccine encoding for oligodendrocyte and myelin-related antigens was employed to attenuate the axonal growth inhibitory properties of myelin in the setting of spinal cord injury. Using a rat spinal cord dorsal hemisection model, the vaccine directed against the inhibitory epitopes of Nogo-A, MAG, OMgp, and TN-R was administered intramuscularly once a week following spinal cord injury, supplemented with local application of specific anti-sera against the four antigens. Anterograde labeling of dorsal column fibers showed active axonal regeneration through the lesion site at the eighth week following the treatment in experimental group but not in control groups. Light microscopic and ultrastructural analysis revealed that vaccination with these myelin-related antigens did not lead to demyelinating disease. OMgp and TN-R levels were down-regulated at the lesion site together with a parallel increase in growth-associated protein 43 levels in the treatment groups. This study reveals the effective approach of a DNA vaccine strategy by attaining the special antibody to direct neutralization of the myelin inhibitors during spinal cord injury.     

氮磷对污水净化中藻类叶绿素含量的影响

在室内模拟生物净化槽中比较研究了氮磷对污水中藻类叶绿素含量和污水净化的影响。污水中磷含量均为７ｍｇ／Ｌ左右,氮含量分别为７７．４、４４．４、２４．８和１３．５ｍｇ／Ｌ,结果发现ＴＮ／ＴＰ＝７７．４／７．０１ｍｇ／Ｌ组,污水经７ｄ净化,藻类叶绿素含量最高,污水净化效果较好。四个实验组比较,叶绿素含量随ＴＮ／ＴＰ比例的上升而上升,呈显著正相关。     

Evaluation of Finnish Diabetes Risk Score in Screening Undiagnosed Diabetes and Prediabetes among U.S. Adults by Gender and Race: NHANES 1999-2010

Objective

To evaluate the performance of Finnish Diabetes Risk Score (FINDRISC) in detecting undiagnosed diabetes and prediabetes among U.S. adults by gender and race.

Methods

This cross-sectional analysis included participants (aged ≥20 years) from the National Health and Nutrition Examination Survey (NHANES) 1999–2010. Sensitivity, specificity, area under the receiver operating characteristic (ROC) curve and the optimal cutoff points for identifying undiagnosed diabetes and prediabetes were calculated for FINDRISC by gender and race/ethnicity.

Results

Among the 20,633 adults (≥20 years), 49.8% were women and 53.0% were non-Hispanic White. The prevalence of undiagnosed diabetes and prediabetes was 4.1% and 35.6%, respectively. FINDRISC was positively associated with the prevalence of diabetes (OR = 1.48 for 1 unit increase, p<0.001) and prediabetes (OR = 1.15 for 1 unit increase, p<0.001). The area under ROC for detecting undiagnosed diabetes was 0.75 for total population, 0.74 for men and 0.78 for women (p = 0.04); 0.76 for White, 0.76 for Black and 0.72 for Hispanics (p = 0.03 for White vs. Hispanics). The area under ROC for detecting prediabetes was 0.67 for total population, 0.66 for men and 0.70 for women (p<0.001); 0.68 for White, 0.67 for Black and 0.65 for Hispanics (p<0.001 for White vs. Hispanics). The optimal cutoff point was 10 (sensitivity = 0.75) for men and 12 (sensitivity = 0.72) for women for detecting undiagnosed diabetes; 9 (sensitivity = 0.61) for men and 10 (sensitivity = 0.69) for women for detecting prediabetes.

Conclusions

FINDRISC is a simple and non-invasive screening tool to identify individuals at high risk for diabetes in the U.S. adults.     

新生BALB/c小鼠肠道潘氏细胞的分布规律

观察BALB/c小鼠小肠潘氏细胞的分布规律。应用石蜡切片、H&E染色技术和LeicaQwin显微图像处理系统,分别对2 d、4 d、6 d、8 d、10 d BALB/c小鼠小肠内潘氏细胞的形态发育、分布规律进行观察和分析。结果发现,4日龄前的BALB/c小鼠各段小肠中肠腺发育尚不完整,未见有潘氏细胞。6日龄后,潘氏细胞开始出现,数量随日龄增加呈递增趋势增长,各日龄间差异显著(P<0.05)。BALB/c小鼠肠道潘氏细胞存在于小肠,十二指肠和空肠较少,回肠较多,各组(不同日龄/肠段)之间差异显著(P<0.05)。     

APRT缺陷型CHO细胞系的建立及其重组蛋白表达能力评价北大核心CSCD

中国仓鼠卵巢(Chinese hamster ovary,CHO)细胞是生产复杂重组药物蛋白的首选宿主细胞,腺嘌呤磷酸核糖转移酶(adenine phosphoribosyltransferase,APRT)催化腺嘌呤与磷酸核糖缩合形成腺苷一磷酸,是嘌呤生物合成步骤中的关键酶。采用基因编辑技术敲除CHO细胞中aprt基因,验证获得的APRT缺陷型CHO细胞系的生物学特性;构建两种真核表达载体:对照载体(含有目的基因增强型绿色荧光蛋白(enhanced green fluorescent protein,EGFP)和弱化载体(含有启动子和起始密码子突变的aprt弱化表达盒及EGFP),分别转染APRT缺陷型和野生型CHO细胞并筛选获得稳定转染的细胞池;重组CHO细胞传代培养60代并用流式细胞术检测EGFP表达的平均荧光强度,并比较不同实验组重组蛋白EGFP的表达稳定性。PCR扩增和测序结果表明,CHO细胞aprt基因成功敲除;获得的APRT缺陷型CHO细胞系在细胞形态、生长增殖、倍增时间等生物学特性方面与野生CHO细胞无显著差异。目的蛋白瞬时表达结果表明,与野生型CHO细胞相比,转染对照载体和弱化载体的APRT缺陷型CHO细胞系中EGFP的表达分别提高了42%±6%和56%±9%;特别是长期传代培养时,转染弱化载体的APRT缺陷型细胞中EGFP表达量显著高于野生型CHO细胞(P<0.05);构建的基于APRT缺陷型CHO细胞系能够明显提高重组蛋白的长期表达稳定性。研究结果为建立高效稳定的CHO细胞表达系统提供了一种有效的细胞工程策略。     

单甲脒农药对模型池塘生态系统群落结构的影响

在３ｍ×１ｍ×１ｍ（Ｖ＝３ｍ３）含有底泥的模型池塘生态系统中研究单脒农药对水生生物群落结构的影晌。规定实验浓度力０、１．５、３．０、６．０和１２．０ｍｇ／Ｌ,每１５ｄ加入１次２５％单甲脒农药水剂,连续加入４次,实验进行２个多月。在实验浓度范围内,童甲脒农药对水生生物群落产生不同程度的影响。浮游生物比较敏感：加药后头几天内,种类、数量及多样性指数下降,浓度越大,影响越明显;大约１周以后,各处理组浮游生物群落逐步得到恢复,实验后期其数量甚至可超过对照水平,但群落结构发生改变,敏感种类少或消失,耐污种类增加,生物多样性降低。底栖生物比较耐污：处理槽大型水生植物的叶绿素含量有所减少,但其种类和生物量未见明显差异;底栖动物种类、数量也未见明显变化。微生物最耐污,在处理槽水层及沉积物中好氧异养菌数量有所增加,沉积物中厌气菌数量也有增加的趋势。青鱼对单甲脒农药较敏感,在１．５ｍｇ／Ｌ以下浓度尚能正常存活和繁殖。单甲脒农药水剂明显增加水体氮、磷含量,尤其磷酸盐含量高,使水体氮、磷比例失调,可能导致水体富营养化。根据综合指标分析,在规定单甲脒盆酸盐浓度＜１．５ｍｇ／Ｌ的实验条件下,水生生物群落结构未见明显改变．